Parkinson's disease (PD) is an insidious neurodegenerative disease that affects more than 1 million people in North America alone. The central aim of this application is to develop insight into the molecular pathways that underlie PD through studies of LRRK2 (leucine-rich repeat kinase 2), the most common genetic cause of the disease. Our central hypothesis is that LRRK2 causes neurodegeneration by transducing signals in the extrinsic apoptotic pathway. Extrinsic apoptotic signaling is critically dependent on FADD (Fas-associated death domain protein) and caspase-8. Preliminary studies in primary neuronal culture indicate that LRRK2-mediated neurodegeneration is FADD- and caspase-8-dependent. Four of five LRRK2 PD mutations enhance LRRK2 oligomerization, which promotes FADD binding to LRRK2 and the subsequent recruitment and activation of caspase-8. Blocking LRRK2 kinase function reverses the enhanced oligomerization, FADD binding and cell death caused by these mutations, supporting a functional relationship between these events. In contrast, one LRRK2 mutation enhances its kinase function, also leading to FADD- and caspase-8-dependent cell death. The physiological relevance of our findings is supported by human post-mortem brain studies that demonstrate activation of the extrinsic apoptotic pathway in LRRK2 patients. Our first specific aim is to test the hypothesis that LRRK2-dependent neurodegeneration requires FADD and caspase-8;we will use RNAi and dominant negative inhibitory proteins in primary neuronal culture. We will also assess whether LRRK2 phosphorylates FADD, and whether it alters other extrinsic pathway downstream signals, including NF-kB and Jun Kinase (JNK). Our second specific aim will test the hypothesis that death domain-containing receptors signal through LRRK2. We will assess whether LRRK2 is a component of receptor-associated death inducing signaling complexes (DISCs), and whether RNAi knockdown of LRRK2 alters the neuronal response to death receptor ligands. Our third specific aim will test the hypothesis that LRRK2 oligomerization is a cell death- related signaling mechanism. We have developed strategies to experimentally increase and decrease LRRK2 oligomerization, and will determine the effect of these manipulations on FADD binding, kinase activity and neurodegeneration. We believe that the results of this work may suggest avenues for exploring the development of rational therapeutic interventions for PD, such as those that block LRRK2 oligomerization or FADD association. PUBLIC HEALTH RELEVANCE: Parkinson's disease is the second commonest neurodegenerative disease of aging, affecting more than 1 million people in North America alone. PD is a progressive disease, and the incidence increases markedly with age, from 20/100,000 overall to 120/100,000 at age 70. Thus, this illness threatens to place a large and increasing burden on our aging society. The central aim of this application is to develop additional insight into the molecular mechanisms that cause PD-related neurodegeneration, through studies of LRRK2, a recently described protein with novel signaling activities. We believe that the results of this work may suggest avenues for exploring the development of rational therapeutic interventions for PD.